Genetic Phenomena Commons^™

Discovery Of Distinct Mechanisms Underlying The Relationship Between Drug Taking And Predisposing Behaviors, Tyler A. Roy

Electronic Theses and Dissertations

Drug addiction is a heritable disease characterized by compulsive drug use. The biological mechanisms driving addiction remain largely unknown.¹ Previous studies show shared genetic mechanisms underlying addiction risk phenotypes such as anxiety, depression, and novelty/sensation seeking.^2,3 Therefore, high-throughput behavioral screening of these traits in single gene knockout mice can allow for the rapid detection of addiction risk candidate genes and mechanisms. Many of these traits are represented in the Knock-Out Mouse Program (KOMP) phenotyping pipeline. Of the initial two hundred twenty-one strains screened in this program, we tested nineteen phenodeviant knock-out mouse strains with C57BL/6NJ ...

Effects Of Nicotinamide Riboside And Beta-Hydroxybutyrate On C. Elegans Lifespan, Jeffery Peters

Undergraduate Honors Theses

The nicotinamide riboside (NR) form of vitamin B₃and the ketone body ß-hydroxybutyrate (BHB) are two of the most promising natural compounds yet identified for the treatment of aging and aging-related diseases. Forms of vitamin B₃are precursors for the synthesis of the coenzymes nicotinamide adenine dinucleotide (NAD(H)) and nicotinamide adenine dinucleotide phosphate (NADP(H)). In aged cells levels of NAD⁺decline, decreasing metabolism and decreasing activity of protective sirtuin protein deacetylases. In aged cells NR, but not more common forms of vitamin B3, boost NAD⁺levels. BHB is naturally produced by the body when individuals fast ...

Flexibility And Constraint In Preimplantation Gene Regulation In Mouse, Colin C. Conine, Marina Krykbaeva, Lina Song, Robert C. Brewster, Nir Friedman, Oliver J. Rando

University of Massachusetts Medical School Faculty Publications

Although many features of embryonic development exhibit remarkable stability in the face of environmental perturbations, it is also clear that some aspects of early embryogenesis can be modulated by non-genetic influences during and after fertilization. Among potential perturbations experienced during reproduction, understanding the consequences of differing ex vivo fertilization methods at a molecular level is imperative for comprehending both the basic biology of early development and the potential consequences of assisted reproduction. Here, we set out to explore stable and flexible aspects of preimplantation gene expression using single-embryo RNA-sequencing of mouse embryos fertilized by natural mating, in vitro fertilization, or ...

Detecting Chromatin Interactions Along And Between Sister Chromatids With Sisterc, Marlies E. Oomen, Adam K. Hedger, Jonathan K. Watts, Job Dekker

University of Massachusetts Medical School Faculty Publications

Accurate chromosome segregation requires chromosome compaction with concordant disentanglement of the two sister chromatids. This process has been studied extensively by microscopy but has remained a challenge for genomic methods, such as Hi-C, because sister chromatids have identical DNA sequences. Here we describe SisterC, a chromosome conformation capture assay that can distinguish interactions between and within sister chromatids. The assay is based on BrdU incorporation during S-phase, which labels the newly replicated strands of the sister chromatids. This is followed by Hi-C, e.g. during different stages of mitosis, and the selective destruction of BrdU containing strands by UV/Hoechst ...

Comprehensive Fitness Maps Of Hsp90 Show Widespread Environmental Dependence, Julia M. Flynn, Ammeret Rossouw, Pamela A. Cote-Hammarlof, Ines Fragata, David Mavor, Carl Hollins Iii, Claudia Bank, Daniel N. Bolon

Open Access Articles

Gene-environment interactions have long been theorized to influence molecular evolution. However, the environmental dependence of most mutations remains unknown. Using deep mutational scanning, we engineered yeast with all 44,604 single codon changes encoding 14,160 amino acid variants in Hsp90 and quantified growth effects under standard conditions and under five stress conditions. To our knowledge, these are the largest determined comprehensive fitness maps of point mutants. The growth of many variants differed between conditions, indicating that environment can have a large impact on Hsp90 evolution. Multiple variants provided growth advantages under individual conditions; however, these variants tended to exhibit ...

Adaptive Evolution Targets A Pirna Precursor Transcription Network, Swapnil Parhad, Tianxiong Yu, Gen Zhang, Nicholas P. Rice, Zhiping Weng, William E. Theurkauf

Open Access Articles

In Drosophila, transposon-silencing piRNAs are derived from heterochromatic clusters and a subset of euchromatic transposon insertions, which are bound by the Rhino-Deadlock-Cutoff complex. The HP1 homolog Rhino binds to Deadlock, which recruits TRF2 to promote non-canonical transcription from both genomic strands. Cuff function is less well understood, but this Rai1 homolog shows hallmarks of adaptive evolution, which can remodel functional interactions within host defense systems. Supporting this hypothesis, Drosophila simulans Cutoff is a dominant-negative allele when expressed in Drosophila melanogaster, in which it traps Deadlock, TRF2, and the conserved transcriptional co-repressor CtBP in stable complexes. Cutoff functions with Rhino and ...

Cell Cycle-Associated Expression Patterns Predict Gene Function In Mycobacteria, Aditya Bandekar, Sishir Subedi, Thomas Ioerger, Christopher M. Sassetti

University of Massachusetts Medical School Faculty Publications

While the major events in prokaryotic cell cycle progression are likely to be coordinated with transcriptional and metabolic changes, these processes remain poorly characterized. Unlike many rapidly-growing bacteria, DNA replication and cell division are temporally-resolved in mycobacteria, making these slow-growing organisms a potentially useful system to investigate the prokaryotic cell cycle. To determine if cell-cycle dependent gene regulation occurs in mycobacteria, we characterized the temporal changes in the transcriptome of synchronously replicating populations of Mycobacterium tuberculosis (Mtb). By enriching for genes that display a sinusoidal expression pattern, we discover 485 genes that oscillate with a period consistent with the cell ...

Targeting Epigenetic Mechanisms To Alleviate Alcoholic Steatosis, Pranoti Mandrekar

Open Access Articles

Alcohol-related liver disease (ALD) is a major health concern and recent studies have reported nearly 1 million alcohol-related deaths from 1999 to 2017 in the United States.¹ ALD is a spectrum of conditions that ranges from early steatosis or fatty liver to inflammation or alcoholic steatohepatitis progressing to fibrosis and cirrhosis. Approximately 8%–20% of alcoholic steatohepatitis patients develop cirrhosis and, in some, alcoholic steatohepatitis can present in the form of acute-on-chronic liver failure, termed alcoholic hepatitis, owing to excessive drinking episodes. Corticosteroids are the first line of therapy for ALD, however, only marginal short-term survival benefit in patients ...

Stride-A Fluorescence Method For Direct, Specific In Situ Detection Of Individual Single- Or Double-Strand Dna Breaks In Fixed Cells, Magdalena M. Kordon, Miroslaw Zarebski, Kamil Solarczyk, Hanhui Ma, Thoru Pederson, Jurek W. Dobrucki

Open Access Articles

We here describe a technique termed STRIDE (SensiTive Recognition of Individual DNA Ends), which enables highly sensitive, specific, direct in situ detection of single- or double-strand DNA breaks (sSTRIDE or dSTRIDE), in nuclei of single cells, using fluorescence microscopy. The sensitivity of STRIDE was tested using a specially developed CRISPR/Cas9 DNA damage induction system, capable of inducing small clusters or individual single- or double-strand breaks. STRIDE exhibits significantly higher sensitivity and specificity of detection of DNA breaks than the commonly used terminal deoxynucleotidyl transferase dUTP nick-end labeling assay or methods based on monitoring of recruitment of repair proteins or ...

Frequent Mutation Of Hypoxia-Related Genes In Persistent Pulmonary Hypertension Of The Newborn, Mingbang Wang, Deyi Zhuang, Mei Mei, Haiyan Ma, Zixiu Li, Fusheng He, Guoqiang Cheng, Guang Lin, Wenhao Zhou

Open Access Articles

AIMS: Persistent pulmonary hypertension of the newborn (PPHN) is characterized by sustained high levels of pulmonary vascular resistance after birth with etiology unclear; Arterial blood oxygen saturation of Tibetan newborns at high latitudes is higher than that of Han newborns at low latitudes, suggesting that genetic adaptation may allow sufficient oxygen to confer Tibetan populations with resistance to pulmonary hypertension; We have previously identified genetic factors related to PPHN through candidate gene sequencing; In this study, we first performed whole exome sequencing in PPHN patients to screen for genetic-related factors.

METHODS AND RESULTS: In this two-phase genetic study, we first ...

Mrna Stem-Loops Can Pause The Ribosome By Hindering A-Site Trna Binding, Chen Bao, Sarah Loerch, Clarence Ling, Andrei A. Korostelev, Nikolaus Grigorieff, Dmitri N. Ermolenko

University of Massachusetts Medical School Faculty Publications

Although the elongating ribosome is an efficient helicase, certain mRNA stem-loop structures are known to impede ribosome movement along mRNA and stimulate programmed ribosome frameshifting via mechanisms that are not well understood. Using biochemical and single-molecule Förster resonance energy transfer (smFRET) experiments, we studied how frameshift-inducing stem-loops from E. coli dnaX mRNA and the gag-pol transcript of Human Immunodeficiency Virus (HIV) perturb translation elongation. We find that upon encountering the ribosome, the stem-loops strongly inhibit A-site tRNA binding and ribosome intersubunit rotation that accompanies translation elongation. Electron cryo-microscopy (cryo-EM) reveals that the HIV stem-loop docks into the A site of ...

Mtorc2-Akt Signaling To Atp-Citrate Lyase Drives Brown Adipogenesis And De Novo Lipogenesis, Camila Martinez Calejman, S. Trefely, S. W. Entwisle, Amelia K. Luciano, Su Myung. Jung, Wen-Yu Hsiao, A. Torres, C. M. Hung, H. Li, N. W. Snyder, J. Villen, K. E. Wellen, David A. Guertin

Open Access Articles

mTORC2 phosphorylates AKT in a hydrophobic motif site that is a biomarker of insulin sensitivity. In brown adipocytes, mTORC2 regulates glucose and lipid metabolism, however the mechanism has been unclear because downstream AKT signaling appears unaffected by mTORC2 loss. Here, by applying immunoblotting, targeted phosphoproteomics and metabolite profiling, we identify ATP-citrate lyase (ACLY) as a distinctly mTORC2-sensitive AKT substrate in brown preadipocytes. mTORC2 appears dispensable for most other AKT actions examined, indicating a previously unappreciated selectivity in mTORC2-AKT signaling. Rescue experiments suggest brown preadipocytes require the mTORC2/AKT/ACLY pathway to induce PPAR-gamma and establish the epigenetic landscape during differentiation ...

A Curated Benchmark Of Enhancer-Gene Interactions For Evaluating Enhancer-Target Gene Prediction Methods, Jill E. Moore, Henry E. Pratt, Michael J. Purcaro, Zhiping Weng

Open Access Articles

BACKGROUND: Many genome-wide collections of candidate cis-regulatory elements (cCREs) have been defined using genomic and epigenomic data, but it remains a major challenge to connect these elements to their target genes.

RESULTS: To facilitate the development of computational methods for predicting target genes, we develop a Benchmark of candidate Enhancer-Gene Interactions (BENGI) by integrating the recently developed Registry of cCREs with experimentally derived genomic interactions. We use BENGI to test several published computational methods for linking enhancers with genes, including signal correlation and the TargetFinder and PEP supervised learning methods. We find that while TargetFinder is the best-performing method, it ...

Reporter Assays For Ebola Virus Nucleoprotein Oligomerization, Virion-Like Particle Budding, And Minigenome Activity Reveal The Importance Of Nucleoprotein Amino Acid Position 111, Aaron E. Lin, William E. Diehl, Yingyun Cai, Courtney L. Finch, Chidiebere Akusobi, Robert N. Kirchdoerfer, Laura Bollinger, Stephen F. Schaffner, Elizabeth A. Brown, Erica Ollmann Saphire, Kristian G. Andersen, Jens H. Kuhn, Jeremy Luban, Pardis C. Sabeti

Open Access Articles

For highly pathogenic viruses, reporter assays that can be rapidly performed are critically needed to identify potentially functional mutations for further study under maximal containment (e.g., biosafety level 4 [BSL-4]). The Ebola virus nucleoprotein (NP) plays multiple essential roles during the viral life cycle, yet few tools exist to study the protein under BSL-2 or equivalent containment. Therefore, we adapted reporter assays to measure NP oligomerization and virion-like particle (VLP) production in live cells and further measured transcription and replication using established minigenome assays. As a proof-of-concept, we examined the NP-R111C substitution, which emerged during the 20132016 Western African ...

Airway Epithelial Specific Deletion Of Jun-N-Terminal Kinase 1 Attenuates Pulmonary Fibrosis In Two Independent Mouse Models, Jos L. Van Der Velden, John F. Alcorn, David G. Chapman, Lennart K. A. Lundblad, Charles G. Irvin, Roger J. Davis, Kelly Butnor, Yvonne M. W. Janssen-Heininger

Open Access Articles

The stress-induced kinase, c-Jun-N-terminal kinase 1 (JNK1) has previously been implicated in the pathogenesis of lung fibrosis. However, the exact cell type(s) wherein JNK1 exerts its pro-fibrotic role(s) remained enigmatic. Herein we demonstrate prominent activation of JNK in bronchial epithelia using the mouse models of bleomycin- or AdTGFbeta1-induced fibrosis. Furthermore, in lung tissues of patients with idiopathic pulmonary fibrosis (IPF), active JNK was observed in various regions including type I and type II pneumocytes and fibroblasts. No JNK activity was observed in adjacent normal tissue or in normal control tissue. To address the role of epithelial JNK1, we ...

Lncrna Rp11-19e11 Is An E2f1 Target Required For Proliferation And Survival Of Basal Breast Cancer, A. Giro-Perafita, L. Luo, A. Khodadadi-Jamayran, M. Thompson, Betul Akgol-Oksuz, A. Tsirigos, B. D. Dynlacht, I. Sanchez, F. J. Esteva

Open Access Articles

Long non-coding RNAs (lncRNAs) play key roles in the regulation of breast cancer initiation and progression. LncRNAs are differentially expressed in breast cancer subtypes. Basal-like breast cancers are generally poorly differentiated tumors, are enriched in embryonic stem cell signatures, lack expression of estrogen receptor, progesterone receptor, and HER2 (triple-negative breast cancer), and show activation of proliferation-associated factors. We hypothesized that lncRNAs are key regulators of basal breast cancers. Using The Cancer Genome Atlas, we identified lncRNAs that are overexpressed in basal tumors compared to other breast cancer subtypes and expressed in at least 10% of patients. Remarkably, we identified lncRNAs ...

Comparing Sensitivity To Change Using The 6-Item Versus The 17-Item Hamilton Depression Rating Scale In The Guided Randomized Controlled Trial, Boadie W. Dunlop, Sagar V. Parikh, Anthony J. Rothschild, Michael E. Thase, Charles Debattista, Charles R. Conway, Brent P. Forester, Francis M. Mondimore, Richard C. Shelton, Matthew Macaluso, Jennifer Logan, Paul Traxler, James Li, Holly Johnson, John F. Greden

Open Access Articles

BACKGROUND: Previous research suggests that the 17-item Hamilton Depression Rating Scale (HAM-D17) is less sensitive in detecting differences between active treatment and placebo for major depressive disorder (MDD) than is the HAM-D6 scale, which focuses on six core depression symptoms. Whether HAM-D6 shows greater sensitivity when comparing two active MDD treatment arms is unknown.

METHODS: This post hoc analysis used data from the intent-to-treat (ITT) cohort (N = 1541) of the Genomics Used to Improve DEpression Decisions (GUIDED) trial, a rater- and patient-blinded randomized controlled trial. GUIDED compared combinatorial pharmacogenomics-guided care with treatment as usual (TAU) in patients with MDD. Percent ...

Comparative Pangenomics Of The Mammalian Gut Commensal Bifidobacterium Longum, Korin Albert, Asha Rani, David Sela

Open Access Articles

Bifidobacterium longum colonizes mammalian gastrointestinal tracts where it could metabolize host-indigestible oligosaccharides. Although B. longum strains are currently segregated into three subspecies that reflect common metabolic capacities and genetic similarity, heterogeneity within subspecies suggests that these taxonomic boundaries may not be completely resolved. To address this, the B. longum pangenome was analyzed from representative strains isolated from a diverse set of sources. As a result, the B. longum pangenome is open and contains almost 17,000 genes, with over 85% of genes found in < /=28 of 191 strains. B. longum genomes share a small core gene set of only ~500 genes, or ~3% of the total pangenome. Although the individual B. longum subspecies pangenomes share similar relative abundances of clusters of orthologous groups, strains show inter- and intrasubspecies differences with respect to carbohydrate utilization gene content and growth phenotypes.

Identification Of The Hyaluronic Acid Pathway As A Therapeutic Target For Facioscapulohumeral Muscular Dystrophy, Alec M. Desimone, John D. Leszyk, Kathryn Wagner, Charles P. Emerson, Jr.

Open Access Articles

Facioscapulohumeral muscular dystrophy (FSHD) is linked to epigenetic derepression of the germline/embryonic transcription factor DUX4 in skeletal muscle. However, the etiology of muscle pathology is not fully understood, as DUX4 misexpression is not tightly correlated with disease severity. Using a DUX4-inducible cell model, we show that multiple DUX4-induced molecular pathologies that have been observed in patient-derived disease models are mediated by the signaling molecule hyaluronic acid (HA), which accumulates following DUX4 induction. These pathologies include formation of RNA granules, FUS aggregation, DNA damage, caspase activation, and cell death. We also observe previously unidentified pathologies including mislocalization of mitochondria and ...

A "Choose-Your-Own" Classroom-Based Activity That Promotes Scientific Inquiry About Rna Interference, Jeremy L. Hsu

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

RNA interference (RNAi), the process that results in the degradation of a target gene’s mRNA, is a fundamental part of eukaryotic gene regulation and is also an important molecular technique that allows for experimental manipulation of gene expression without altering DNA sequences. Despite the importance of RNAi, there have been relatively few lecture-based activities designed to teach about the consequences of this process and counter common misconceptions. I present here an inquiry-based activity that is centered around a “choose your own experiment” design where students generate hypotheses and critically evaluate their ideas by choosing several simulated experiments. The activity ...